The laboratory has been focused during the last year on immune-mediated tumor cell destruction. Certain lymphocytes like natural killer (NK) cells and cytotoxic T cells (CTL) have the ability to lyse virally-infected and certain tumor cells. There are two proposed major mechanisms whereby they accomplish this cytotoxic effect (1) the exocytosis of lytic granules and (2) the expression of cytotoxic molecules like tumor necrosis factor (TNF) or a related molecule known as Fas-ligand. The cytolytic granules contain a variety of unique proteins, including a pore-forming protein (perforin) and a family of serine protease enzymes (granzymes). We identified, sequenced and cloned a novel member of the human granzyme family which cleaves synthetic substrates after the amino acids arginine or lysine and therefore has a "tryptase" activity. We designated the enzyme HNK-Tryp-2. The gene was a single copy gene which we localized ( in collaboration with a group in Australia) to human chromosome 5, in close proximity to another granzyme tryptaseA called granzyme A. The HNK-Tryp-2 is only expressed in lymphoid cells and seems to be completely restricted in its expression to NK cells and activated T cells. We are currently trying to define the biological properties of this enzyme. In addition, using a murine renal cancer model Renca, we are investigating the role of immune-mediated effects on apoptosis of tumor cells. Apoptosis can be induced in the Renca cells in vitro by combinations of interferon gamma and tumor necrosis factor, or antibodies which cross-link the Fas molecule on the surface of the tumor cells. The protein synthesis inhibitor cycloheximide substantially enhances these apoptotic effects. We are currently investigating whether such effects play an important role in immune-mediated rejection of Renca in vivo.